Pectin polymers are important constituents of plant cell walls. Pectin is a hetero-polysaccharide with a backbone composed of alternating homogalacturonan (smooth regions) and rhamnogalacturonan (hairy regions). The smooth regions are linear polymers of 1,4-linked alpha-D-galacturonic acid. The galacturonic acid residues can be methyl-esterified on the carboxyl group to a varying degree, usually in a non-random fashion with blocks of polygalacturonic acid being completely methyl-esterified.
Pectinases can be classified according to their preferential substrate, highly methyl-esterified pectin or low methyl-esterified pectin and polygalacturonic acid (pectate), and their reaction mechanism, beta-elimination or hydrolysis. Pectinases can be mainly endo-acting, cutting the polymer at random sites within the chain to give a mixture of oligomers, or they may be exo-acting, attacking from one end of the polymer and producing monomers or dimers. Several pectinase activities acting on the smooth regions of pectin are included in the classification of enzymes provided by the Enzyme Nomenclature (1992) such as polymethylgalacturonase (EC 4.2.2.2), pectin lyase (EC 4.2.2.10), polygalacturonase (EC 3.2.1.15), exo-polygalacturonase (EC 3.2.1.67), exo-polygalacturonate lyase (EC 4.2.2.9) and exo-poly-alpha-galacturonosidase (EC 3.2.1.82).
Glycosyl hydrolases are classified into families according to their three-dimensional structure or folding; conventionally the Clustal W method is used the for family determination. Based on amino acid sequence alignment and the Clustal W method, a polypeptide or protein can be classified into a specific glycosyl hydrolase family, ie either a known family or a novel and hitherto unknown family (The Sanger Centre: Protein Families Database of alignments and HMMs; www. sanger.ac.uk). At present known polymethylgalacturonases belong to family 28 of glycosyl hydrolases (ExPASy--molecular biology WWW server of the Swiss Institute of Bioinformatics (SIB)).
Polymethylgalacturonases have been cloned from various microbial organisms. However, up till very recently all polymethylgalacturonases were known as requiring divalent cations for maximum activity, calcium ions being the most stimulatory. In contrast hereto, Japanese patent application Kokai 10-313858 discloses a novel polymethylgalacturonase of family 28 which is believed not to require the presence of divalent cations for maximum activity, ie this novel enzyme is capable of exerting its action in an aqueous solution which further comprises calcium chelators. Calcium chelators are known to be present in a number of industrial processes including scouring of cotton.
It is an object of the present invention to provide improved industrial enzymatic processes which are more efficient and/or have higher cost-benefit than the hitherto known processes.